Viruses cause various diseases, and particularly, a typical one among pathogenic viruses that become a problem in the field of stockbreeding is Avian influenza virus. Avian influenza virus belongs to the Orthomyxoviridae family, and causes much damage to poultry, such as hens and turkey. Avian influenza viruses are classified into highly pathogenic avian viruses, low pathogenic avian viruses and non-pathogenic avian viruses according to their pathogenicity. The highly pathogenic avian viruses are classified as “List A species” by the Office International des Epizootics (OIE) and as class I infectious livestock diseases in Korea.
The influenza viruses are classified into three types, A, B and C, according to the antigenic properties of their matrix proteins and nucleocapsid proteins. Moreover, according to the differences in the antigenic structures of haemagglutinin (HA), which assists in host cell receptor binding and the fusion between the host cell membrane and the viral envelop to cause a virus infection, and neuraminidase (NA) which plays an important role when the viruses bud from cells after proliferation, the influenza viruses are further classified into 16 HA and 9 NA subtypes, each. Theoretically, 144 kinds of virus subtypes could exist by the combination of the two proteins.
Infection with Avian influenza virus occurs mainly by direct contact with avian excreta and also spreads by droplets, water, human feet, feeding cars, instruments, devices, feces attached to the outer surface of eggs, and the like. In the symptoms of the viral infection, respiratory symptoms, diarrhea and a rapid decrease in egg production are commonly shown, although the symptoms vary depending on the pathogenicity of infected viruses. In some cases, head portions, such as crests, show cyanosis, and sometimes edema appears on the face or feathers flock together at one point. Mortality caused by the viral infection varies from 0% to 100% depending on the viral pathogenicity. The viral infection requires precise diagnosis because its symptoms are similar to those of other diseases such as Newcastle disease, infectious laryngotracheitis, mycoplasma infections, and the like.
About 23 outbreaks of highly pathogenic avian influenza have been recorded worldwide during 1959-2003, but were mostly localized events. Outbreaks of H5N1 subtype high-pathogenic avian influenza that occurred in Korea in December, 2003 occurred in more than 30 countries, including Europe, Africa and most countries in Southeast Asia such as Japan, China, Thailand, Vietnam and Indonesia, thus becoming pandemic.
Although it is known that humans cannot be infected by avian influenza, prevention of avian influenza is being of paramount importance to public health sector due to the case of human infection with H5N1 in Hongkong in 1997, isolation of H9N2 avian influenza viruses from humans in 1999 and human cases of H7 avian influenza infection in Canada in 2004. According to a report of the World Health Organization (WHO) (http://www.who.int/csr/disease/avian_influenza/country/cases_table—2006—06—20/en/index.html), it was confirmed that the 228 persons were infected with H5N1 subtype virus in 10 countries, and 130 persons of them died during a period ranging from 2003 to Jun. 20, 2006. In Korea, since low-pathogenic avian influenza by H9N2 subtype had occurred in 1996, it reoccurred in 1999.
When avian influenza outbreaks occur, most countries respond by killing all of the infected animals involved in the outbreak, and countries experiencing outbreaks cannot export poultry products. Accordingly, avian influenza viruses can be regarded as being among primary factors that interfere with the development of the livestock industry. Furthermore, when there is a risk of human infection, the damage spread to a wide range of industries, including the tourist industry and the transport industry.
Recently, considerable efforts have been made worldwide to develop anti-viral agents. Commercially available anti-viral agents include lamibudine that is used for the treatment of HIV (Human Immunodeficiency Virus)-1 and hepatitis B, gancyclovir that is used for the treatment of herpes virus infections, and ribavirin that is used mainly for the treatment of symptoms of respiratory syncytial virus infection but can be used for the treatment of symptoms of various virus infections in emergency. In addition, zanamivir RELENZA™ and oseltamivir, TAMIFLU™ which are synthesized artificially as neuraminidase inhibitors of influenza virus are also commercially available.
However, the use of amantadine and its analogue, rimantadine, which were approved for treatment of influenza virus A, has been limited due to the appearance of resistant virus and its side effect. Recently, virus resistant to oseltamivir among H5N1 avian influenza viruses appeared, and thus, the development of various anti-virus agents is urgently required.
Meanwhile, Alnus japonica is a deciduous tree belonging to the genus Alnus of the Betulaceae and is commonly called an Alnus japonica tree. About 30 species of Alnus japonica are distributed in the Northern Hemisphere and the South America, and about 9 species of Alnus japonica are distributed in Korea. It grows near swampy areas, its height is about 20 m and its bark is of a deep purplish-brown color. Its winter bud is a long oval shape just like the shape of an egg turned upside down, which has three ridge lines and a peduncle. The leaves of Alnus japonica grow alternately, and they are oval shaped, egg-shaped or lanceolate. Both sides of the leaf are lustrous and leaf margins are saw-toothed. The flower of Alnus japonica blooms in March to April, is unisexual and forms a catkin. Staminate spike bears staminate flower and each bract has 3-4 flowers. There are four perianths and four stamens in each flower. The fruit ripens in October and 2-6 fruits are produced. It is long egg-shaped and looks like a pine cone.
Meanwhile, triterpenoid-based compounds contain α-amyrin, α-amyrin acetate, baurenol acetate, β-amyrin, β-amyrin acetate, daturaolone germanicol acetate, lupeol acetate, Lup-20(29)-en-3-one, olean-18-en-3-one, and taraxasterol, and sesquiterpenoids include 11,13-α-dehydroglucozaluzanin C, 10-α-hydroxy-8-dseoxyglucosid, 8-epideacylcynaropicrin, 8-epideacylcynaropicrin glucoside, glucozaluzanin C ixerin, picriside B and the like (M. Tamai et al., Planta Med., 1989; S. Seo et al., J. Am. Chem. Soc., 1981; T. Akihisa et al., Phytochemistry, 1994; W. Kisiel, Phytochemistry, 1992; H. Fuchino et al., Chem. Pharm. Bull., 1995; K. Shiojima et al., Chem. Pharm. Bull., 1996; A. Hisham et al., Phytochemistry, 1995).
In Korean Patent Registration Nos. 10-0721703 and 10-0769050, the present inventors confirmed the antiviral activity of Alnus japonica extracts. However, the Alnus japonica extracts have limited use, because they have a shortcoming in that they show antiviral activity only when they are administered at high concentrations.
Accordingly, the present inventors have made many efforts to develop a natural material, which has low toxicity to normal cells and shows an excellent effect of inhibiting viral proliferation even when it is administered at low concentrations. As a result, the present inventors found that triterpenoid-based compounds or diaryl hepatonoid-based compounds extracted from Alnus japonica show an excellent effect of inhibiting avian influenza virus activity, thereby completing the present invention.